Article

Selective capability of SynCAM and neuroligin for functional synapse assembly

Center for Basic Neuroscience, The University of Texas Southwestern Medical Center, Dallas, Texas 75390-9111, USA.
The Journal of Neuroscience : The Official Journal of the Society for Neuroscience (Impact Factor: 6.34). 02/2005; 25(1):260-70. DOI: 10.1523/JNEUROSCI.3165-04.2005
Source: PubMed

ABSTRACT

Synaptic cell adhesion is central for synapse formation and function. Recently, the synaptic cell adhesion molecules neuroligin 1 (NL1) and SynCAM were shown to induce presynaptic differentiation in cocultured neurons when expressed in a non-neuronal cell. However, it is uncertain how similar the resulting artificial synapses are to regular synapses. Are these molecules isofunctional, or do all neuronal cell adhesion molecules nonspecifically activate synapse formation? To address these questions, we analyzed the properties of artificial synapses induced by NL1 and SynCAM, compared the actions of these molecules with those of other neuronal cell adhesion molecules, and examined the functional effects of NL1 and SynCAM overexpression in neurons. We found that only NL1 and SynCAM specifically induced presynaptic differentiation in cocultured neurons. The induced nerve terminals were capable of both spontaneous and evoked neurotransmitter release, suggesting that a full secretory apparatus was assembled. By all measures, SynCAM- and NL1-induced artificial synapses were identical. Overexpression in neurons demonstrated that only SynCAM, but not NL1, increased synaptic function in immature developing excitatory neurons after 8 d in vitro. Tests of chimeric molecules revealed that the dominant-positive effect of SynCAM on synaptic function in developing neurons was mediated by its intracellular cytoplasmic tail. Interestingly, morphological analysis of neurons overexpressing SynCAM or NL1 showed the opposite of the predictions from electrophysiological results. In this case, only NL1 increased the synapse number, suggesting a role for NL1 in morphological synapse induction. These results suggest that both NL1 and SynCAM act similarly and specifically in artificial synapse induction but that this process does not reflect a shared physiological function of these molecules.

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Available from: Deniz Atasoy, Aug 21, 2014
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    • "binant GABA A R or GlyR isoform of interest . Co - culture approaches have previously been developed to understand the roles of synaptic adhesion molecules ( including neurexin and neuroligin ) in the formation of glutamatergic or GABAergic synapses ( Scheiffele et al . , 2000 ; Biederer et al . , 2002 ; Dean et al . , 2003 ; Graf et al . , 2004 ; Sara et al . , 2005 ; Kim et al . , 2006 ; Dong et al . , 2007 ; Fuchs et al . , 2013 ) or to investigate the impact of disease - causing neuroligin mutations on GABAergic synaptogenesis ( Chubykin et al . , 2005 ; Sun et al . , 2011 ) . They have also been employed to characterize the functional properties of inhibitory post - synaptic currents ( IPSCs ) ,"
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    ABSTRACT: Fast inhibitory neurotransmission in the brain is mediated by wide range of GABAA receptor (GABAAR) and glycine receptor (GlyR) isoforms, each with different physiological and pharmacological properties. Because multiple isoforms are expressed simultaneously in most neurons, it is difficult to define the properties of inhibitory postsynaptic currents mediated by individual isoforms in vivo. Although recombinant expression systems permit the expression of individual isoforms in isolation, they require exogenous agonist application which cannot mimic the dynamic neurotransmitter profile characteristic of native synapses. We describe a neuron-HEK293 cell co-culture technique for generating inhibitory synapses incorporating defined combinations of GABAAR or GlyR subunits. Primary neuronal cultures, prepared from embryonic rat cerebral cortex or spinal cord, are used to provide presynaptic GABAergic and glycinergic terminals, respectively. When the cultures are mature, HEK293 cells expressing the subunits of interest plus neuroligin 2A are plated onto the neurons, which rapidly form synapses onto HEK293 cells. Patch clamp electrophysiology is then used to analyze the physiological and pharmacological properties of the inhibitory postsynaptic currents mediated by the recombinant receptors. The method is suitable for investigating the kinetic properties or the effects of drugs on inhibitory postsynaptic currents mediated by defined GABAAR or GlyR isoforms of interest, the effects of hereditary disease mutations on the formation and function of both types of synapses, and synaptogenesis and synaptic clustering mechanisms. The entire cell preparation procedure takes 2 – 5 weeks.
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    • "In contrast, NL2 and NL3 KOs caused selective impairments in subsets of GABAergic synapses (Chubykin et al., 2007; Gibson et al., 2009; Poulopoulos et al., 2009; Etherton et al., 2011; Fö ldy et al., 2013; Rothwell et al., 2014). Overexpression of all neuroligin isoforms, conversely, increased synapse numbers as assessed morphologically (Boucard et al., 2005; Chih et al., 2005; Ko et al., 2009b; Sara et al., 2005; Zhang et al., 2009). In addition, overexpression of NL1 enhanced both NMDAR-and AMPAR-mediated excitatory postsynaptic currents (EPSCs), overexpression of NL2 selectively increased inhibitory postsynaptic currents (IPSCs), and overexpression of NL4 paradoxically decreased NMDAR-and AMPAR-mediated EPSCs, whereas overexpression of NL3 produced no electrophysiological effect (Chubykin et al., 2007; Ko et al., 2009b; Zhang et al., 2009; Chanda et al., 2013). "
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    ABSTRACT: Neuroligins are postsynaptic cell-adhesion molecules that bind presynaptic neurexins and are genetically linked to autism. Neuroligins are proposed to organize synaptogenesis and/or synaptic transmission, but no systematic analysis of neuroligins in a defined circuit is available. Here, we show that conditional deletion of all neuroligins in cerebellar Purkinje cells caused loss of distal climbing-fiber synapses and weakened climbing-fiber but not parallel-fiber synapses, consistent with alternative use of neuroligins and cerebellins as neurexin ligands for the excitatory climbing-fiber versus parallel-fiber synapses. Moreover, deletion of neuroligins increased the size of inhibitory basket/stellate-cell synapses but simultaneously severely impaired their function. Multiple neuroligin isoforms differentially contributed to climbing-fiber and basket/stellate-cell synapse functions, such that inhibitory synapse-specific neuroligin-2 was unexpectedly essential for maintaining normal climbing-fiber synapse numbers. Using systematic analyses of all neuroligins in a defined neural circuit, our data thus show that neuroligins differentially contribute to various Purkinje-cell synapses in the cerebellum in vivo. Copyright © 2015 Elsevier Inc. All rights reserved.
    Full-text · Article · Aug 2015 · Neuron
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    • " disease ( Saura et al . , 2011 ; Bie et al . , 2014 ) . These cell - adhesion molecules are critical for synapse specification and function via interactions with partners that constitute the neurotransmission machinery on both sides of the synapse ( Scheiffele et al . , 2000 ; Missler et al . , 2003 ; Graf et al . , 2004 ; Prange et al . , 2004 ; Sara et al . , 2005 ; Gerrow et al . , 2006 ; Chubykin et al . , 2007 ; Mukherjee et al . , 2008 ) . NL1 is specific to excitatory synapses where it promotes the retention of α - amino - 3 - hydroxyl - 5 - methyl - 4 - isoxazole - propionate receptors ( AMPARs ; Heine et al . , 2008 ; Mondin et al . , 2011 ) as well as the clustering of N - methyl - D - asp"
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    Full-text · Article · Jul 2015 · Frontiers in Synaptic Neuroscience
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